Alcoholic liver disease (ALD) is one of the most common liver diseases worldwide characterized by the accumulation of lipids within the liver, inflammation and the possibility of progressing to cirrhosis and liver failure. More importantly, there are currently no effective treatments for ALD and liver transplantation remains the only therapeutic option for end stage liver disease. Previous studies have shown that ALD is a result of a combination of endoplasmic reticulum (ER) stress, lipid metabolism dysregulation and inflammation. It has been previously reported that alcohol disrupts gut microbiota homeostasis and causes increased endotoxins that contribute to the pathology of ALD. However, the detailed mechanism(s) underlying ALD and disease progression is poorly understood. We have discovered that sphingosine kinase 2 (SphK2) deficient (SphK2-/-) mice on an alcohol diet exhibit increased steatosis and inflammation compared to wild type mice. Sphingosine 1-phosphate receptor 2 (S1PR2) and SphK2 have been previously shown to play a key role in nutrient metabolism and signaling. However, their roles in alcohol-induced liver injury have not been characterized. The overall objective of this project is to determine the molecular mechanism(s) by which disruption of S1PR2- mediated SphK2 signaling contributes to ALD. Aim 1. First, we will determine the role of S1PR2 and SphK2 in alcohol-induced liver injury. We will examine the effects of alcohol on primary hepatocytes and Kupffer cells derived from S1PR2 deficient (S1PR2-/-) and SphK2-/- mice. We will examine various pathways and mechanisms of injury including hepatic lipid metabolism dysregulation, ER stress and inflammation. For in vivo studies, we will adopt the acute on chronic alcohol mouse model from NIAAA that recapitulates the drinking pattern of human alcoholic liver disease patients to study the effects of S1PR2 and SphK2 deficiency in ALD. We will further characterize the expression patterns of S1PR2 and SphK2 in human ALD liver samples. Aim 2. Second, we will identify potential mechanisms by which S1PR2 and SphK2 protect against alcohol-induced liver injury. We will examine various cellular stress pathways and the role of S1PR2 and SphK2 in regulating inflammatory mediators. Finally, we will evaluate the therapeutic potential of targeting the S1PR2/SphK2- mediated signaling pathway using an S1PR2 chemical agonist CYM-5520 to attenuate alcohol-induced liver injury. Accomplishing these aims could provide important information on the development of effective treatments and drug targets against ALD.